Trypanosomes are parasites that cause African trypanosomiasis or "sleeping sickness". They reside in the host animal's bloodstream and evade its immune system by sequentially expressing different surface proteins, called variable surface glycoproteins or VSGs. Earlier work showed that (i) the trypanosome genome contains a battery of VSG genes, only one of which is expressed at a time and (ii) DNA rearrangements occur in the vicinity of the VSG genes. The objective of this research is to determine the molecular mechanism(s) at the DNA and RNA level that are responsible for this antigenic variation. Using recombinant DNA techniques we have constructed cDNA and genomic DNA libraries of six different trypanosome clones, three T. bruccei and three T. rhodesiense. We plan to determine the VSG cDNA sequences and the corresponding chromosomal gene sequences (and flanking regions) isolated from genomic libraries of trypanosomes that are (i) expressing those VSG genes and (ii) not expressing those VSG genes. We expect these sequences will contribute as strongly to understanding trypanosome antigenic variation as similar sequencing efforts have contributed to understanding the genetic basis of immunoglobulin diversity. We also plan to search for clusters of VSG genes in the trypanosome genome by "chromosome walking" using the genomic libraries and to experimentally estimate the number of VSG genes available to a trypanosome. We then plan to use this VSG gene structural analysis as a foundation for designing assays to characterize and purify the enzyme system(s) responsible for the switch in expression of one VSG gene to another. And finally we hope that this information will provide insights into means of inhibiting this switch in a way that will prevent the antigenic variation of trypanosomes.